The proposed studies are concerned with the mechanism of activation of fluorenylamides and related carcinogens by cultured embryonic cells of the rat and human. We intend to use rat-embryo cells of low passage, a system recently developed in this laboratory, for assays of the carcinogenic potential of arylamides and of their activated derivatives. This culture system will also be applied to the growth of human-embryo cells in vitro. The following metabolic reactions will be investigated: 1) N-hydroxylation, a reaction obligatory for carcinogenesis in the rat that has not yet been demonstrated in human cells. 2) O-Acetylation and O-sulfation of the arylhydroxamic acids which arise in the N-hydroxylation of arylamides. O-Acetates and O-Sulfates are regarded as ultimate agents in hepatocarcinogenesis in the rat. However, there is as yet no evidence for their occurence or for their role in carcinogenesis of the extrahepatic tissues which are transformed following administration of the respective arylamides or arylhydroxamic acids. 3) One-electron oxidation of arylhydroxamic acids to free radicals. 4) Intramolecular transacetylation of arylhydroxamic acids to O-acyl-N-arylhydroxylamines. Although it has been postulated that this reaction is implicated in carcinogenesis by arylamides, direct evidence for the importance of this pathway in carcinogenesis is lacking. Syntheses as well as assays in vitro of the transforming potential of nitroxyl radicals and of O-acyl-N-arylhydroxylamines are proposed for the unequivocal identification of these hypothetical carcinogens and for he determination of their role in carcinogenesis. Finally, the enzymatic degradation of certain sugar nucleotides by normal and transformed cells of the rat and human will be explored. These studies are undertaken in the search of a biochemical alteration generally characteristic for and indicative of malignant transformation.